In the first three-year period 31P and 17O NMR methods were used to study the stereochemistry of adenylyl transfer reactions. As a continuation in the stereochemistry involving phosphorus, we will focus on the reactions catalyzed by phospholipases A1, A2, C and D in this three-year period. The phospholipids labeled with 17O and 18O (chiral at phosphorus) will be synthesized and theri configurations will be determined by various spectroscopic techniques. These compounds will be used as substrates for phospholipase C and phospholipase D. The [16O, 17O, 18O] phosphomonoesters resulted from enzymatic hydrolysis will be analyzed for its configuration by the method of Knowles which is based on the 18O isotopes shift developed by Cohn and the 17O quadrupolar effect in 31P NMR developed in the first three-year period of this project. Thiophospholipids (sulfur analogues of phospholipids) will be synthesized, resolved in diastereomers, and analyzed for its configuration. The stereospecificity of phospholipases A1, A2, C and D toward the diastereomers will be determined. The stereospecificity in the enzyme-lipid interaction will also be determined for the phospholipase A2. As a secondary aim, 2H and 14N NMR of thiophospholipids in the gel phase will be studied to explore the conformation of the phosphate head group in phospholipids.